Autosomal dominant polycystic kidney disease (ADPKD) is one of the most common life-threatening genetic diseases, and is a leading cause of renal failure. The majority of cases are caused by mutations in the PKD1 gene, which codes for polycystin-1, PC1, a protein predicted to function as a cell adhesion protein mediating cell-cell and cell matrix interactions. This large multi-modular protein contains 16 copies of a novel immunoglobulin (Ig)-like fold, the polycystic kidney disease (PKD) domain. Tandem repeats of Ig-like domains are a common feature of proteins with structural and mechanical roles, such as titin (an elastomeric protein of muscle), fibronectin (an elastic extracellular matrix protein component) or cadherin (a mechano-chemical cell adhesion protein). Recent single molecule experiments have shown that such proteins are highly extensible and elastic. [unreadable] [unreadable] We propose that PC1 functions mechanically by providing a flexible and elastic linkage between cells. The mechanical properties of PC1 may be vital for maintaining the architectural integrity of the kidney. Mutations may alter PC1's cell adhesion and mechanical properties and lead to the alterations in cell-cell interactions and abnormal tissue development which are characteristic of ADPKD. However, there is very little information about the structural organization of the extracellular domain of PC1, or how mechanical forces may affect its structural features. Neither is it known how mutations might alter PC1's structure and mechanical properties. [unreadable] [unreadable] In this R21 project we plan to use a novel combination of single molecule and protein engineering tools to test whether PC1 is a highly elastic and extensible molecule, as predicted from its structure. In Aim 1 we will use single molecule force spectroscopy to examine the stability and mechanical properties of the diverse protein domains found in PC1. In Aim 2 we will examine the effects of naturally occurring mutations on the mechanical stability and unfolding kinetics of PC1's PKD domains. The long-term goal of our research is to gain a better understanding of the function of normal and mutant forms of PC1 and lay the foundation for therapeutic approaches to the currently untreatable ADPKD. [unreadable] [unreadable]